APOBEC3H structure reveals an unusual mechanism of interaction with duplex RNA

Bohn, Jennifer; Thummar, Keyur; York, Ashley; Raymond, Alice; Brown, William Clay; Bieniasz, Paul D.; Hatziioannou, Théodora; Smith, Janet L.

doi:10.1038/s41467-017-01309-6

Cited by 53 publications

(138 citation statements)

References 57 publications

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“…Since A3H and A3A are singledomain enzymes with nearly identical cores, we constructed additional chimeras to ask what surface residues are required to endow A3A with A3H properties. We first tested other components of the RNA binding interface because our work implicated loop 1 in nucleolar retention (see above) and structural studies showed select loop 1 residues together with loop 7 or ␣-helix 6 residues combining to bind duplex RNA (40,41,48). We therefore generated A3A chimeras that exchanged loop 7 or ␣-helix 6 alone or in combination with loop 1 and assessed localization (structural overlay in Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Furthermore, interactions between several A3s and cellular proteins can be disrupted by RNase A treatment, suggesting that RNA plays a pivotal role in regulating A3 activities. This is further evidenced by recent studies requiring RNase A treatment in order to purify high quantities of recombinant A3H protein from cell lysates (40,41,48).…”

mentioning

confidence: 94%

“…Multiple groups recently succeeded in solving the X-ray crystal structure of A3H and discovered a surprising RNA-mediated dimerization mechanism (40,48). In these structures, two A3H monomers dimerize around a short, 7-to 9-base-pair RNA duplex.…”

mentioning

confidence: 99%

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APOBEC3H Subcellular Localization Determinants Define Zipcode for Targeting HIV-1 for Restriction

Salamango

Becker

McCann

et al. 2018

Molecular and Cellular Biology

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APOBEC enzymes are DNA cytosine deaminases that normally serve as virus restriction factors, but several members, including APOBEC3H, also contribute to cancer mutagenesis. Despite their importance in multiple fields, little is known about cellular processes that regulate these DNA mutating enzymes. We show that APOBEC3H exists in two distinct subcellular compartments, cytoplasm and nucleolus, and that the structural determinants for each mechanism are genetically separable. First, native and fluorescently tagged APOBEC3Hs localize to these two compartments in multiple cell types. Second, a series of genetic, pharmacologic, and cell biological studies demonstrate active cytoplasmic and nucleolar retention mechanisms, whereas nuclear import and export occur through passive diffusion. Third, APOBEC3H cytoplasmic retention determinants relocalize APOBEC3A from a passive cell-wide state to the cytosol and, additionally, endow potent HIV-1 restriction activity. These results indicate that APOBEC3H has a structural zipcode for subcellular localization and selecting viral substrates for restriction.

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Section: Resultsmentioning

confidence: 99%

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confidence: 94%

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APOBEC3H Subcellular Localization Determinants Define Zipcode for Targeting HIV-1 for Restriction

Salamango

Becker

McCann

et al. 2018

Molecular and Cellular Biology

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“…The first ever A3H structure was recently published of a macaque A3H (macA3H) bound to dsRNA [Fig. (e)] . This was also the first structure of an APOBEC bound to RNA, albeit of unknown sequence (resolution was not high enough to identify the nucleotides of copurified RNA).…”

Section: Introductionmentioning

confidence: 88%

APOBEC3s: DNA‐editing human cytidine deaminases

Silvas

Schiffer

2019

Protein Science

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Nucleic acid editing enzymes are essential components of the human immune system that lethally mutate viral pathogens and somatically mutate immunoglobulins. Among these enzymes are cytidine deaminases of the apolipoprotein B mRNA editing enzyme, catalytic polypeptide‐like (APOBEC) super family, each with unique target sequence specificity and subcellular localization. We focus on the DNA‐editing APOBEC3 enzymes that have recently attracted attention because of their involvement in cancer and potential in gene‐editing applications. We review and compare the crystal structures of APOBEC3 (A3) domains, binding interactions with DNA, substrate specificity, and activity. Recent crystal structures of A3A and A3G bound to ssDNA have provided insights into substrate binding and specificity determinants of these enzymes. Still many unknowns remain regarding potential cooperativity, nucleic acid interactions, and systematic quantification of substrate preference of many APOBEC3s, which are needed to better characterize the biological functions and consequences of misregulation of these gene editors.

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“…The activity of A3H is improved through its ability to oligomerize, dimerization is uniquely mediated through an RNA intermediate [130][131][132]. Interestingly, two studies recently identified that A3H may be able to dimerize through an RNA mediated mechanism that involves loops 1 and 7 as well as helix 6 and that the presence of bound RNA alters the activity of the A3H [131,133]. These findings warrant future work on the exact nature of A3H oligomerization and activity.…”

Section: Deamination-dependent Restriction Of Hiv By A3hmentioning

confidence: 95%

Biochemical Basis of APOBEC3 Deoxycytidine Deaminase Activity on Diverse DNA Substrates

2018

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Apolipoprotein B mRNA-editing enzyme-catalytic, polypeptide-like 3 (APOBEC3) enzymes are a family of single-stranded (ss)DNA cytosine deaminases that serve as a host restriction factors for retrotransposons and viruses that contain a ssDNA intermediate. While the APOBEC3 family was originally expanded to restrict endogenous retroelements, the majority of these targets are now inactivated and the family has been found to act on different targets, such as viral ssDNA as a mechanism of viral defense. Some of the family members also catalyze "offtarget" deaminations in human ssDNA and have been implicated in somatic mutagenesis that can lead to cancer.My PhD thesis research investigated the biochemical mechanisms underlying both the benefits and the risks of the A3 family of enzymes. The central hypothesis of this work is that A3 enzymes have evolved distinct biochemical mechanisms to deaminate ssDNA that differentiate A3 restriction of retroelements and retroviruses from A3-induced somatic mutagenesis.Therefore, we investigated the biochemical mechanisms the A3 enzymes utilize during restriction of Human Immunodeficiency Virus (HIV) in both deamination-dependent and deaminationindependent modes, as well as the unique mechanisms employed during mutation of genomic DNA. There are seven APOBEC3 members (A-H, excluding E) and of these, four members (A3D, A3F, A3G, A3H) have been identified to restrict HIV replication in CD4 T+ cells, and currently three members (A3A, A3B and A3H haplotype I) have been implicated in somatic mutagenesis.The APOBEC3 enzymes that restrict HIV replication function optimally in the absence of the HIV viral infectivity factor (Vif). Vif targets APOBEC3 for degradation via the proteasome in HIV infected cells. For APOBEC3s that are able to fortuitously escape Vif mediated degradation and become encapsidated, the enzymes can deaminate cytosines to form uracils in viral (-)DNA. Replication of (-)DNA to (+)DNA causes HIV-1 reverse transcriptase to incorporate adenines opposite uracils which creates C/G→T/A transition mutations. While restriction of HIV most commonly occurs through this deamination-dependent mechanism, APOBEC3s have also been identified to interfere with HIV reverse transcriptase processes in a deamination-independent manner, although this mechanism is secondary to the deaminationdependent mode. In order to restrict retroelements and viruses such as HIV, the A3 enzymes iii require an efficient processive ssDNA scanning mechanism that allows them to search for, and locate cytosines on ssDNA in the finite amount of time the substrate is available during formation of the double-stranded DNA provirus. To understand if this requirement was lost in A3 enzymes unable to restrict HIV, we studied A3C. Human A3C is not able to restrict HIV, in comparison to the more active gorilla and chimpanzee A3C orthologs that can restrict HIV, however an explanation for this difference was lacking. A polymorphism, S188I, in human A3C, which is found in less than 3% of the global population lead...

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APOBEC3H structure reveals an unusual mechanism of interaction with duplex RNA

Cited by 53 publications

References 57 publications

APOBEC3H Subcellular Localization Determinants Define Zipcode for Targeting HIV-1 for Restriction

APOBEC3H Subcellular Localization Determinants Define Zipcode for Targeting HIV-1 for Restriction

APOBEC3s: DNA‐editing human cytidine deaminases

Biochemical Basis of APOBEC3 Deoxycytidine Deaminase Activity on Diverse DNA Substrates

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